Biocatalytic reactions are currently primarily performed in systems which are aqueous or at least contain a considerable amount of water. This mode of procedure is disadvantageous if hydrophobic substrates which are poorly water-soluble or non-water-soluble, e.g. steroids, lipids, hydrocarbons, long-chain aliphatic or aromatic alcohols, aldehydes or carboxylic acids, are to be reacted by biocatalysis or if an enzyme-catalyzed synthesis is to be performed as a reversal of a hydrolysis reaction. In addition, enzymes themselves can also be strongly hydrophobic. In aqueous systems, this can cause agglomeration and, consequently, a loss of biocatalytic activity.
The procedures known and employed in the past for the biocatalytic reaction of hydrophobic substrates are:
1. The use of enzymes in organic solvents or water-containing solvent mixtures. PA1 2. The use of two-phase and multiphase systems. PA1 3. The use of one-phase, surfactant-containing systems, especially of reverse micellar phases.
In systems of Type 1, there is a danger of enzyme denaturation. This can only be retarded by means of the selection of suitable solvents and by enzyme immobilization. The residence times of the catalysts are therefore only low. The use of microorganisms and other cells as biocatalysts is possible only to a limited extent.
There is also the danger, in liquid systems of Type 2, of denaturation at the phase boundary. Moreover, the reaction speed is low due to the low substrate concentration in the aqueous phase. An advantageous variant of Type 2 is the use of hydrophilic polymer gels for enzyme inclusion. These polymer gels, e.g. based on gelatin, agar or alginate, can also be used in organic solvents. However, the concentration of the hydrophobic substrate in the gel is low because of the hydrophilic character of the polymer gel. The result is a low conversion rate in biocatalysis.
Systems of Type 3 exhibit more advantageous properties. Reverse micelles are globular surfactant aggregates which can be formed by surfactants in organic solvents, e.g. hexane, heptane, cyclohexane, benzene or dodecanol in the presence of slight amounts of water.
The water forms a "waterpool" inside the micelle in which enzymes can be included or solubilized. It has been demonstrated frequently that the enzyme activity can be maintained by this means for a considerable time. In addition, it is known that microorganisms, e.g. E. coli or B. subtilis, can maintain a part of their enzyme activity in such phases. Hydrophobic substrates can generally be introduced directly into the organic solvent in the case of reverse micellar systems.
However, preparative steps for retaining the surfactant aggregates and for separating the products are necessary in the processing mode, which imposes considerable expense. An improvement is possible in a continuous processing mode using immobilized reverse micelles. A hollow fiber membrane system has been suggested for this purpose. These membranes add significant expense, are often not resistant to solvents, tend to become clogged and represent a considerable flow resistance, so that the process must be carried out under pressure.
In general, it must be concluded that one-phase, surfactant-containing systems are extremely unreliable for industrial biocatalysis since, in particular, the problems of a simple separation of the product, of the removal of surfactant impurities from the product and of the continuous processing mode remain as yet unsolved.